Seyedi Fatemeh, Farsinejad Alireza, Nematollahi-Mahani Seyed Noureddin
Department of Anatomy, School of Medicine, Jiroft University of Medical Sciences, Jiroft, Iran.
Stem Cell Research Lab., Afzalipour School of Medicine, Kerman University of Medical Sciences, Kerman, Iran.
Tissue Cell. 2017 Apr;49(2 Pt B):227-232. doi: 10.1016/j.tice.2017.03.001. Epub 2017 Mar 8.
Tissue engineering is a new strategy which proposed to treat numerous human diseases nowadays. Three dimensional (3D) scaffolds fill the gap between two dimensional cell culture (2D) and animal tissues through mimicking the environmental behaviors surrounding the cells. In this study, hUCMs into insulin producing cells in fibrin scaffold were differentiated compare to conventional culture condition. Differentiation rate was estimated by real time PCR, immunocytochemistry (ICC) and the chemiluminesence (CLIA) and enzyme immunoassay (EIA). Real time PCR's results showed an increasing expression in NKX2.2, PDX1 and INS (producing the hormone insulin) genes in fibrin scaffold. Furthermore ICC analysis exhibited that insulin and pro-insulin proteins were more in fibrin scaffolds. CLIA and EIA on insulin and C peptide secretion indicated that both of groups were sensitive to the glucose challenge test but significant higher response was observed in fibrin scaffold (6.5 fold in 3D, 1.8 fold in 2D culture). It could be concluded that differentiation of hUCM cells into insulin producing cells in fibrin scaffold 3D culture system is much more efficient than 2D conventional culture system.
组织工程是目前提出的一种治疗多种人类疾病的新策略。三维(3D)支架通过模拟细胞周围的环境行为,填补了二维细胞培养(2D)与动物组织之间的空白。在本研究中,与传统培养条件相比,人脐带间充质干细胞(hUCMs)在纤维蛋白支架中被诱导分化为胰岛素分泌细胞。通过实时定量聚合酶链反应(real time PCR)、免疫细胞化学(ICC)、化学发光免疫分析(CLIA)和酶免疫分析(EIA)来评估分化率。实时定量聚合酶链反应结果显示,在纤维蛋白支架中,NKX2.2、PDX1和INS(产生胰岛素激素)基因的表达增加。此外,免疫细胞化学分析表明,纤维蛋白支架中胰岛素和胰岛素原蛋白含量更多。对胰岛素和C肽分泌的化学发光免疫分析和酶免疫分析表明,两组对葡萄糖激发试验均敏感,但在纤维蛋白支架中观察到显著更高的反应(3D培养中为6.5倍,2D培养中为1.8倍)。可以得出结论,在纤维蛋白支架3D培养系统中,hUCM细胞向胰岛素分泌细胞的分化比2D传统培养系统更有效。